PAD4 inhibition modulates inflammatory responses in macrophages during Mycobacterium tuberculosis infection

INTRODUCTION: Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), is the leading cause of death due to an infectious disease  worldwide. During infection, macrophages actively participate in the immune response and release inflammatory mediators involved in the control of infection. In addition, they can release macrophage extracellular traps (METs), a process that can involve the activity of the enzyme peptidylarginine deiminase 4 (PAD4), which citrulinates proteins. However, the role of PAD4 in the regulation of macrophage effector functions during Mtb infection remains poorly understood. OBJECTIVE: To evaluate the effect of PAD4 inhibition, using the inhibitors GSK484 and Cl-amidine at different concentrations, on the response of macrophages to Mtb infection. METHODOLOGY: Bone marrow–derived macrophages from WT mice were infected in vitro with Mtb and stimulated or not with IFN-γ in the presence or absence of the PAD4 inhibitors Cl-amidine or GSK484. After 24 hours, IL-1β production was measured by ELISA and the expression of Arg1, iNOS, and MHC II was analyzed by flow cytometry. CEUA 7711141123 RESULTS: Compared to controls, PAD4 inhibition with Cl-amidine significantly reduced IL-1β production at 10 µM, while no differences were observed at concentrations of 50 and 100 µM. In IFN-γ-stimulated BMDMs, Cl-amidine treatment significantly reduced IL-1β production at 10, 50, and 100 µM compared to untreated cells. By flow cytometry, we found that PAD4 inhibition with Cl-amidine (10 µM) in IFN-γ-stimulated Mtb-infected macrophages induced increased expression of Arg1, iNOS, and MHC II. CONCLUSION: Preliminary data indicate that PAD4 inhibition modulates the response of macrophages to Mtb infection in the presence and absence of IFN-γ stimulation, altering IL-1β production and parameters associated with cellular activation. Additional studies are underway to investigate this phenotype and the underlying mechanisms. CAPES process: 88887.271252/2026-00